Abstract: The brain is an extraordinarily complex network made of billions of neurons. The synapse, the contact point between neurons, has been the centre of interest for decades. Starting with Ram[unreadable]n Cajal, scientists have been borrowing tools from other disciplines to study this tiny, but crucial structure. The breakthrough truly came after the introduction of electrophysiology, which allows the measurement of neuronal activity at an extremely high signal-to-noise ratio. It became clear that synaptic transmission is conducted via the presynaptic release of neurotransmitters from synaptic vesicles and the subsequent activation of postsynaptic receptors. So far, changes in synaptic transmission have generally been attributed to the modification of postsynaptic receptors. Nevertheless, newly emerged evidence has demonstrated that the receptors do not act autonomously and synaptic vesicles indeed make significant contribution to synaptic plasticity. Despite great interest and effort, the lack of a direct and precise measurement obstructs further investigation of synaptic vesicles and thus impedes a complete comprehension of such mutual modulation. Borrowing from nanotechnology, I devised a single-vesicle imaging approach using quantum dot (Qdot), which for the first time provides superior spatiotemporal resolution matching or even exceeding that of patch-clamp recording. With this original tool, I unveiled the existence, kinetics and regulatory mechanisms of an unconventional mode of vesicle reuse in mammalian central synapses. Here, I propose that synaptic modulation is composed of a sequence of coordinated and equally profound changes at both pre- and postsynaptic terminals. To test it, I will develop ""smart"" nanosensors made of Qdots and aptamers, a new class of synthetic oligonucleic acids. These nanosensors can precisely target designated synaptic structures or proteins and simultaneously report orchestrated pre- and postsynaptic changes in situ. To demonstrate the power of this revolutionary tool, I will deploy it to address some fundamental and long-standing questions in synaptic physiology, such as the coupling of vesicle reacidification and refilling after retrieval, the fluctuation of neurotransmitter concentration around the synaptic cleft, and the acculturation of presynaptic vesicles and postsynaptic receptors. Because Qdots are ideal for in vivo imaging, I will screen for aptamers that can deliver these nanosensors to designated synapses in the intact brain such that synaptic transmission can be studied in defined neuronal circuits. The success of this project promises a quantum leap in our understanding of neurotransmission. The technical and scientific advances made from this work can be readily transplanted to other biomedical fields because vesicles turnover and surface receptor trafficking are widely involved in almost all biological processes. Moreover, both Qdots and aptamers are modular units that can be linked with all kinds of diagnostic or therapeutic molecules. Therefore, this project has a broad impact beyond neuroscience. . Public Health Relevance: Synapse is the most essential element for transmitting information in brain neuronal network which is responsible for our cognition and daily behavior. By inventing and deploying multifunctional nanoporbes, we will investigate the most fundamental aspects of synaptic transmission including synaptic vesicles, neurotransmitters and receptors in mammalian central nervous system. As these components of synapses evidently became abnormal in most neurological disorders, our finding will bridge the gap between basic and clinic research for better understanding, detection, characterization, and treatment of neurological disorders.